Cherry Red tobacco is the superior variant showing red dapples in cured leaves due to the demethylation of nicotine to nornicotine during maturation and curing. Fungi is known for its capacity of converting nicotine to nornicotine. However, to the best of our knowledge, the endophytic community of Cherry Red tobacco is never reported. Here we sampled mature leaves from Cherry Red and ordinary tobacco at lower, centre and upper plant sections, and analysed the internal transcribed spacer region (ITS) diversity using high-throughput sequencing. Results revealed a significantly different fungal community of foliar endophyte in Cherry Red and ordinary tobacco. In comparison to the ordinary control, a higher diversity and co-occurrence network complex was found in Cherry Red samples, especially in the centre and upper leaves, where the red dapples had mainly emerged. More taxa were enriched in the Cherry Red than ordinary tobacco leaves at all plant sections. Particularly, Aspergillus, a fungus that has been demonstrated to efficiently convert nicotine to nornicotine, was specifically enriched in upper Cherry Red leaves, which presented the most red dapples after curing. A less robust network structure was detected in the Cherry Red tobacco compared to the ordinary tobacco, corresponding to the vulnerability of red dapple production to agronomic and environmental disturbances. The nearest taxon index and β nearest taxon index indicated that the local community structuration of tobacco endophytic fungi was mainly driven by deterministic process, while the community turnover among plant sections was stochastic. In conclusion, our study provides the earliest information of the endophytic fungal community in Cherry Red tobacco leaf; the community diversity, composition and network features synchronously varied with the appearance of red dapples, suggesting their potential role in the formation of Cherry Red tobacco.